The present invention relates to a method of inducing oligodendrocyte precursor cells (OPCs) through direct reprogramming from human somatic cells into which a nucleic acid molecule encoding an Oct4 protein has been introduced or Oct4 protein-treated human somatic cells.
Due to the progression of an aging society, personalized cell therapeutic agents for healthier aging and a healthy life without disease are medications essential for improving the quality of life. Multiple sclerosis known as a disease of the central nervous system is a demyelinating disease whose cause is unknown, and in severe cases, is accompanied by sensory and motor impairment. However, there is no fundamental therapy, except treatment with medications for reducing symptoms. Accordingly, transplantation of OPCs (OPC) that can differentiate into oligodendrocytes enabling the generation of myelin sheaths has attracted attention as a main therapeutic method, and research to obtain cells which will be used for the transplantation is progressing using embryonic stem cells and adult stem cells.
Embryonic stem cells are pluripotent cells which are able to differentiate into all types of human cells having the ability to divide indefinitely, unlike somatic cells. Adult stem cells are multipotent cells which are able to be extracted from a patient, and as a representative example, neural stem cells (NSCs) are well known. NSCs, which are adult stem cells, can overcome immune rejection in the treatment of neurological diseases, and therefore has attracted attention as a cell therapeutic agent. However, NSCs are not effective because they have a considerably low ability to differentiate into oligodendrocytes, and are limited in number of cells because they should be obtained from a patient's own cerebral tissue. Embryonic stem cells also have disadvantages to be overcome for clinical use. First, there is an ethical issue because it is necessary to destroy a fertilized embryo to obtain embryonic stem cells, and when cells differentiated from the embryonic stem cells are transplanted into a patient, immune rejection occurs.
Among various methods attempting to overcome such problems, a method of dedifferentiation from differentiated cells to undifferentiated cells has attracted attention, and dedifferentiation encompasses the generation of pluripotent stem cells such as embryonic stem cells using differentiated cells. After induced pluripotent stem cells (iPS cells) were developed through gene introduction by Prof. Shinya Yamanaka, Japan in 2006, a variety of studies for applying such cells to a therapeutic agent are progressing.
After the report in which stem cells having similar characteristics to embryonic stem cells are established when four genes (dedifferentiation-inducible factors; Oct4, Sox2, c-Myc and Klf4) were introduced into mouse or human somatic cells, and then cultured under embryonic stem cell culture conditions for a long time (Cell 126:663-676, 2006; Science 318:1917-1920, 2007) had been suggested, various methods capable of replacing genes for clinical use have been studied. However, because of still insufficient results of the study on human somatic cells, difficulty in defining a dedifferentiation-inducing mechanism, and a risk of forming teratoma, it is difficult to apply iPS cells to clinical trials. A direct reprogramming method is a method which has been recently suggested as an alternative for such iPS cells, and includes two types of techniques for introducing genes specifically expressed in cells and inducing the cells to desired cells without the pluripotency stage through the regulation of a growth signal by combination of dedifferentiation-inducible factors and low molecular weight substances. Such a method is highly appreciated in that it has only advantages of various stem cells, and eliminates many of the factors which inhibit a clinical use.
In recent years, the possibility of establishment of mouse OPCs has been shown by the Marius Wernig research team using three genes (Olig2, Sox10, Zfp536) and by the Paul J Tesar research team using three genes (Olig2, Sox10, NKX6.2) in the United States, and it has been reported that they made a success of direct reprogramming from mouse somatic cells to OPCs. However, there is no still report in which human somatic cells are used. In addition, the Shengding and Mickie Bhatia research teams in the United States demonstrated the establishment of neural stem cells through direct reprogramming by introducing an Oct4 gene into a human somatic cell, and thus suggested a new paradigm in which the Oct4 gene can regulate the expression of nervous system-related genes in cells.
Therefore, the inventors intensively attempted to induce OPCs from human somatic cells through direct reprogramming, resulting in confirming the possibility of induction of OPCs by introducing Oct4 into human somatic cells and treating several low molecular weight substances involved in forming oligodendrocytes, and thus completed the present invention.